Summary of Work: Work from our laboratory demonstrates that P450 monooxygenases of the CYP2J subfamily metabolize arachidonic acid (AA) to epoxyeicosatrienoic acids (EETs) and hydroxy-eicosatetraenoic acids (HETEs). These eicosanoids possess potent biological activities including effects on vascular/airway smooth muscle tone, ion transport, and peptide hormone secretion. Current research involves: (1) characterization of the CYP2J subfamily P450s at the molecular and biochemical levels; (2) studies on the regulation of CYP2J gene expression; (3) evaluation of the functional roles that CYP2J products play in cell/organ physiology; and (4) examination of this enzyme pathway in animal models of disease. Our laboratory has cloned the cDNAs to four different members of the mammalian CYP2J subfamily (human CYP2J2, rat CYP2J3, mouse Cyp2j5 and mouse Cyp2j6). The recombinant CYP2J proteins expressed in Sf9 insect cells with baculovirus possess distinct enzymological properties. CYP2J2 metabolizes AA to EETs primarily; CYP2J3 catalyzes the formation of EETs and 19-HETE; and Cyp2j5 metabolizes AA to EETs and several mid-chain HETEs. Northern analysis and protein immunoblotting reveals that the CYP2J mRNAs and proteins are widely distributed in mammalian tissues. CYP2J2 and CYP2J3 are expressed at highest levels in the heart but are also present in the liver, intestine, lung, kidney, pituitary/hypothalamus, and pancreas. Cyp2j5 mRNA is expressed primarily in the kidney and at lower levels in liver. Cyp2j6 transcripts are expressed at high levels in intestine and at lower levels in the heart, lung, kidney, liver, and hypothalamus. The levels of CYP2J mRNAs and proteins are unaltered by xenobiotics (e.g. phenobarbital, aromatic hydrocarbons, acetone, and clofibrate) and physiologic manipulations (e.g. increased dietary salt, hyperoxic exposure, fasting- refeeding). Synthetic EETs were shown to affect LHRH secretion in GT1 hypothalamic neuronal cells, modulate K+ transport in GH4C1 pituitary cells, and influence Cl- transport in lung epithelial cells. The human CYP2J2 gene mapped to Chr. 1p31.3-p31.2 by FISH. Cyp2j5 and Cyp2j6 mapped to the corresponding region of mouse Chr.4. The mouse Cyp2j5 gene was cloned and sequenced to determine intron/exon boundaries.